Interleukin-12 (IL-12), formerly known as cytotoxic lymphocyte maturation factor (CLMF) or natural killer cell stimulatory factor (NKSF), is a cytokine that has pleiotropic activities including stimulation of the proliferation of activated T and NK cells (1, 2), induction of INF-.gamma. production by peripheral blood mononuclear cells (2, 3), and enhancement of the lytic activity of NK/LAK cells (2, 4).
IL-12 is a heterodimeric molecule with an approximate molecular weight of about 75 kDa consisting of two disulfide-linked subunits: p35, having an approximate molecular weight of about 35 kDa, and p40, having an approximate molecular weight of about 40 kDa, (2, 4-6). The p40 subunit shares amino acid sequence homology with the interleukin-6 receptor (IL-6R) (7) and therefore belongs to the cytokine receptor superfamily, whereas p35 has a distant but significant relationship to the IL-6/G-CSF cytokine family (8). It has been speculated that the p35/p40 heterodimer could represent a cytokine (p35) and soluble cytokine receptor (p40) complex, with the cellular IL-12 receptor providing function analogous to the IL-6 signal transducing protein, gp130 (7, 8).
The biological activity of IL-12 is mediated by the binding of the intact IL-12 molecule to plasma membrane receptors on activated T or NK cells (9,10); however, the contributions of the individual subunits to receptor binding and signal transduction remain unknown. Studies with neutralizing antibodies to human IL-12 (11) and site-specific chemical modification (12) suggested that the p40 subunit contains epitopes important for IL-12 binding to its receptor. Also, studies with human/mouse chimeric molecules indicated that p35 is responsible for the species specificity of the heterodimer for biological activities (13).
We investigated both the binding and biological activities of each IL-12 subunit. COS cells transfected with only the p40 cDNA produced both p40 monomer and p40 homodimer having an approximate molecular weight of about 80 kDa, the latter capable of binding to the IL-12 receptor but unable to mediate cellular proliferation. The 80 kDa p40 homodimer acts as a receptor antagonist useful in regulating the biological activity of IL-12 in immune responses.